Exonuclease VII repairs quinolone-induced damage by resolving DNA gyrase cleavage complexes

Abstract

The widely used quinolone antibiotics act by trapping prokaryotic type IIA topoisomerases, resulting in irreversible topoisomerase cleavage complexes (TOPcc). Whereas the excision repair pathways of TOPcc in eukaryotes have been extensively studied, it is not known whether equivalent repair pathways for prokaryotic TOPcc exist. By combining genetic, biochemical, and molecular biology approaches, we demonstrate that exonuclease VII (ExoVII) excises quinolone-induced trapped DNA gyrase, an essential prokaryotic type IIA topoisomerase. We show that ExoVII repairs trapped type IIA TOPcc and that ExoVII displays tyrosyl nuclease activity for the tyrosyl-DNA linkage on the 5′-DNA overhangs corresponding to trapped type IIA TOPcc. ExoVII-deficient bacteria fail to remove trapped DNA gyrase, consistent with their hypersensitivity to quinolones. We also identify an ExoVII inhibitor that synergizes with the antimicrobial activity of quinolones, including in quinolone-resistant bacterial strains, further demonstrating the functional importance of ExoVII for the repair of type IIA TOPcc.

Mutational Analysis of Quinolone-Resistant Determining Region gyrA and parC Genes in Quinolone-Resistant ESBL-Producing E. Coli

“…This finding would suggest the other mode of mechanisms may also confer a high resistance but at a very low rate. 8 In terms of antibiotic susceptibility profile, all the QR ESBL-E. coli isolates were also MDR bacteria. These pathogens, however, were susceptible to carbapenems despite the low level of resistance (0.7%) to ertapenem (a member of carbapenems).…”

“…Thus, changes such as single nucleotide mutation of the genes may confer resistance to quinolone drugs by the bacterial cells. 8,9 Genetic mechanisms of quinolone resistance may also be mediated by plasmid bearing one or multiple genes such as qnrA, qnrB, and qnrC. 10 These genes encode for proteins that protect DNA gyrase and topoisomerase IV from quinolone inhibition.…”

“…Studies have shown that the quinolone-resistant bacteria may confer varying degrees of resistance either chromosomally or plasmidmediated or in combination. 8,9 Among these resistance mechanisms, target-mediated resistance caused by specific mutations in the genes coding for subunits of DNA gyrase (gyrA) and topoisomerase IV (parC), is commonly reported worldwide. [11][12][13][14] These resistance mutations often occur in a region called the quinolone resistance determining region (QRDR) in the encoded gene, which is in close proximity to the amino-terminal domain.…”

Rapid, DNA-induced interface swapping by DNA gyrase

“…In eukaryotic cells, pathways exist for the removal of the covalently bound topoisomerase II enzyme and exposure of the DNA lesion to regular DNA repair mechanisms (Pommier, Huang et al 2014). In bacterial cells such pathways are not well known (but see (Huang, Michaels et al 2021)).…”

“…In eukaryotic cells, pathways exist for the removal of the covalently bound topoisomerase II enzyme and exposure of the DNA lesion to regular DNA repair mechanisms (Pommier, Huang et al 2014). In bacterial cells such pathways are not well known (but see (Huang, Michaels et al 2021)). Some work has also suggested that in Escherichia coli cleavage complexes can induce illegitimate recombination between two DNA segments (Ikeda, Moriya et al 1981, Ikeda 1994, Ikeda, Shiraishi et al 2004).…”